Zhong J, Douglas A L, Hatch T P
Department of Molecular Sciences, University of Tennessee, Memphis, TN 38163, USA.
Mol Microbiol. 2001 Jul;41(2):451-62. doi: 10.1046/j.1365-2958.2001.02531.x.
Chlamydiae are bacterial parasites that carry out a distinct developmental cycle within host cells; however, the mechanisms by which these organisms regulate stage-specific gene expression are not known. We identified a DNA element located between nucleotide (nt) -135 and -90 upstream from the transcription start point of the late stage-specific CRP operon (omcAB) of Chlamydia trachomatis, to which a protein in extracts of chlamydiae harvested at 23 h after infection binds. A recombinant protein of C. trachomatis open reading frame (ORF) CT267, which is homologous to bacterial integration host factor (IHF) and the heat-unstable nucleoid protein (HU), bound to the same element and produced the same DNase I footprint as the protein in chlamydial extracts. Recombinant ORF CT267 protein bound with high affinity to the DNA element and induced a sharp bend in a DNA fragment containing the binding site, suggesting that ORF CT267 encodes a protein with IHF-like activity, and recombinant protein had a positive effect on in vitro transcription of the CRP operon. IHF-binding activity and IHF protein were detected in extracts of C. trachomatis during the early to intermediate phases of the late stage of the developmental cycle (between 17 and 30 h after infection), but were absent in the extreme late phase of the cycle and in the infectious form of chlamydiae. The presence of an IHF binding site upstream of the CRP operon and the presence of chlamydial IHF-like protein when late stage genes are transcribed suggests that the chlamydial IHF may play a role in stage-specific gene expression.
衣原体是在宿主细胞内进行独特发育周期的细菌寄生虫；然而，这些生物体调节阶段特异性基因表达的机制尚不清楚。我们在沙眼衣原体晚期特异性CRP操纵子（omcAB）转录起始点上游核苷酸（nt）-135至-90之间鉴定出一个DNA元件，感染后23小时收获的衣原体提取物中的一种蛋白质与之结合。沙眼衣原体开放阅读框（ORF）CT267的重组蛋白，与细菌整合宿主因子（IHF）和热不稳定类核蛋白（HU）同源，与同一元件结合，并产生与衣原体提取物中的蛋白质相同的DNase I足迹。重组ORF CT267蛋白与该DNA元件高亲和力结合，并在含有结合位点的DNA片段中诱导急剧弯曲，表明ORF CT267编码一种具有IHF样活性的蛋白质，且重组蛋白对CRP操纵子的体外转录有积极作用。在发育周期晚期的早期至中期阶段（感染后17至30小时之间），在沙眼衣原体提取物中检测到IHF结合活性和IHF蛋白，但在周期的极晚期阶段和衣原体的感染形式中不存在。CRP操纵子上游存在IHF结合位点，以及在转录晚期基因时存在衣原体IHF样蛋白，这表明衣原体IHF可能在阶段特异性基因表达中发挥作用。
